Phosphoinositide signaling at the excitatory synapse generates Ca2+ signals that modify the function of the synapse, and propagate to the nucleus where they result in transcriptional responses. The Snyder/Worley project will examine three critical stages in signal transduction at the synapse. Group 1 metabotropic receptors (mGluR) stimulate phospholipase C to generate IP3, which evokes release of calcium from intracellular pools. Recent studies indicate that mGluR5 is essential for behavioral responses to cocaine. The Worley laboratory has studied mGluRs and their functional modulation by the immediate early gene Homer. In ongoing studies, they find that mice with a genetic deletion of Homer 2 exhibit enhanced responses to cocaine . These observations focus attention on the role of Homer in regulating mGluR function in models of drug addiction. Aim 1 will generate mouse genetic models with mutations of mGluR that disrupt specific interactions with Homer and Shank. Aim 2 will examine a novel signaling/scaffold protein for mGluR. A family of proteins, termed Hopi, was identified based on their coupling to mGluR by interacting with Homer. Preliminary studies indicate that Hopi couples group 1 mGluR to activation of p21-activated kinase (Pak) and perhaps CDK5. Studies will include generation and analysis of conditional Hopi knockout mice, and as well as molecular studies of signaling. Aim 3 will examine molecular mechanisms that target IP3 receptors to the excitatory synapse. The Snyder laboratory has a long-standing interest in the brain receptor for inositol trisphosphate (IP3R) and has defined important interactions involved in its signal transduction. The approach exploits a recent observation that Shank induces the formation of spines that include IP3R. In order to define regions of the IP3R that are essential for accumulation in spines, IP3R deletion mutants will be co-expressed with Shank in neurons. Critical finding will be confirmed in slice cultures. Parallel biochemical studies will identify proteins that interact with the domain of the IP3R that is essential in its spine localization and function. These studies will provide important new insights into how the mGluR5/IP3R signaling complex is assembled at the synapse, and how perturbations contribute to mental illness.